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CHORI-239: Chinese Pangolin (Manis pentadactyla) BAC Library |
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The CHORI-239 “Chinese Pangolin” BAC library has been constructed by Baoli Zhu, Ph.D., at the INstitute of Microbiology, Chinese Academy of Sciences, Beijing. The library has been arrayed in 384-well microtiter dishes and was subsequently characterized by Boudewijn ten Hallers, in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. The preparation of the library followed the general cloning approach developed in our laboratory Osoegawa et al., 1998. Frozen tissues from a Chinese Pangolin was kindly provided by Drs.Wenui Nie and Bing Su from the Kunming Institute of Zoology, The Chinese Academy of Sciences. The tissue were originally part of the Kunming Cell Bank of Type Culture Collection. High-molecular-weight DNA isolated from tissue turned into powder under liquid nitrogen using a pestel and mortar. The powder was extracted to create a chromatine suspension and the chromatine bound DNA was embedded in agarose blocks and treated with a combinations of proteinase K and SDS. The embedded DNA was partially digested with a combination of EcoRI restriction enzyme and EcoRI methylase then size fractionated by pulsed-field electrophoresis. DNA fragments from the appropriate size fraction were cloned into the pBAC-GMR vector between the two EcoRI sites. The ligation products were transformed into DH10B (T1 resistant) electro-competent cells (Invitrogen). The library has been arrayed into plates 590 384-well microtiter dishes and has subsequently been gridded onto 13 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct BAC clones, stamped in duplicate. BAC library construction was funded by NIH grant HG025323-03 under the auspices of the NIH-funded BAC Resource Network. Data for CHORI-239 Chinese Pangolin BAC Library:
Segment | 1 | 2 | All | Vector | pTARBAC2.1 | pTARBAC2.1 | pTARBAC2.1 | Restriction Enzyme | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | DNA Source | kidney | kidney | kidney | Plate Numbers | 1-288 | 289-528 | 1-528 | Plate Count | 288 | 240 | 528 | Empty Wells | N/A | N/A | N/A | Non-Recombinant Clones | N/A | N/A | N/A | Non-Insert Clones | N/A | N/A | N/A | Recombinant Clones | 110592 | 92160 | 202752 | Average Insert Size | N/A | N/A | N/A | Genomic Coverage | N/A | N/A | N/A |
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